A Soluble Form of the MHC Class I-Specific CD160 Receptor Is Released from Human Activated NK Lymphocytes and Inhibits Cell-Mediated Cytotoxicity

Abstract

CD160 is a GPI-anchored lymphocyte surface receptor in which expression is mostly restricted to the highly cytotoxic CD56dimCD16+ peripheral blood NK subset. We previously reported that MHC class I (MHC-I) molecules bind to CD160 receptors on circulating NK lymphocytes and that this interaction triggers their cytotoxic activity and cytokine production. We also observed that CD160 surface expression on NK cells is down-modulated upon activation with PMA or IL-2. In this study, we further report that short-time incubation of NK lymphocytes with IL-15 converts the membrane-bound CD160 to a soluble form through a proteolytic cleavage involving a metalloprotease. Thus, CD160 is no longer detected at the cell surface, but can be immunoprecipitated from the NK cell culture medium. Interestingly, CD160 transcript remains highly expressed during the process of protein shedding. In addition, we demonstrate that CD160 mRNA synthesis can be induced in CD56bright separated lymphocytes following exposure to IL-15. By producing a Flag-tagged soluble CD160 protein, we establish that its binding to MHC-I molecules results in the inhibition of the cytotoxic CD8+ T lymphocyte activity and of the CD160-mediated NK cell cytotoxicity. Thus, we show that activated NK lymphocytes release a soluble form of CD160 that functionally impairs the MHC-I-specific cytotoxic CD8+ T lymphocyte responsiveness.

Alternative mRNA splicing creates transcripts encoding soluble proteins from most LILR genes

“…These soluble molecules may arise by proteolytic cleavage from the cell surface by matrix metalloproteinases [31][32][33][34]. For example, a soluble form of CD160 shed from the surface of CD56 dim CD16 1 NK cells, upon IL-15 stimulation, inhibits CD8…”

“…In addition to the range of molecular interactions sLILRB1 may influence, it has the potential to modulate the response of a variety of cell types in view of the broad distribution of LILRB1 expression. However, the influence of sLILRB1 on immune interaction is likely to be localised to the source cell, where it will be concentrated.The negative regulation of membrane-associated immunoreceptors by soluble isoforms has been described previously (e. g. [34,48]). CTLA-4 is a member of the immunoglobulin superfamily that encodes both a membrane-associated and soluble protein due to alternative transcription.…”

“…T-cell and NK-cell cytotoxicity, presumably by competitive binding of the MHC class I ligand [34]. Soluble isoforms of membrane-associated proteins can also result from alternatively spliced mRNA transcripts that lack TM encoding regions, for example LILRB4 (ILT3/LIR5 [3]) or the Fas molecule [35].…”

Identification of CD245 as myosin 18A, a receptor for surfactant A: A novel pathway for activating human NK lymphocytes

“…PB-NK cell purity was shown to be >95% as assessed by flow cytometry. The YT2C2 NK cell line, the murine mastocytoma cell line P815, and the Burkitt-lymphoma B-cell line Raji (all purchased from the ATCC, Manassas, USA), the Epstein-Barr Virus (EBV)-infected B cell line (locally produced 13 ) were cultured in Roswell Park Memorial Institute (RPMI) 1640 medium supplemented with 1% penicillin/streptomycin, 2 mML-glutamine, and 10% heat-inactivated fetal calf serum (FCS) (Perbio Science, Villebon-sur-Yvette, France).…”

“…13 Briefly, 2 to 10 £ 10 6 cells or 100 mg of tissue were lysed in 1% Nonidet-P40 or Brij58 lysis buffer (Sigma) for 1 h at 4 C. After centrifugation at 10,000 rpm and 4 C, post-nuclear lysates were subjected to immunoprecipitation with either DY12 or control IgG mAb and protein G-Sepharose beads. The precipitated proteins were separated by SDS-8% PAGE and transferred onto a nitrocellulose membrane (Millipore, Bedford, USA).…”